Dual Transcriptional Profiling of Carrot and 'Candidatus Liberibacter solanacearum' at Different Stages of Infection Suggests Complex Host-Pathogen Interaction.

The interactions between the phloem-limited pathogen 'Candidatus Liberibacter solanacearum' haplotype C and carrot (Daucus carota subsp. sativus) were studied at 4, 5, and 9 weeks postinoculation (wpi), by combining dual RNA-Seq results with data on bacterial colonization and observations of the plant phenotype. In the infected plants, genes involved in jasmonate biosynthesis, salicylate signaling, pathogen-associated molecular pattern- and effector-triggered immunity, and production of pathogenesis-related proteins were up-regulated. At 4 wpi, terpenoid synthesis-related genes were up-regulated, presumably as a response to the psyllid feeding, whereas at 5 and 9 wpi, genes involved in both the terpenoid and flavonoid production were down-regulated and phenylpropanoid genes were up-regulated. Chloroplast-related gene expression was down-regulated, in concordance with the observed yellowing of the infected plant leaves. Both the RNA-Seq data and electron microscopy suggested callose accumulation in the infected phloem vessels, likely to impair the transport of photosynthates, while phloem regeneration was suggested by the formation of new sieve cells and the upregulation of cell wall-related gene expression. The 'Ca. L. solanacearum' genes involved in replication, transcription, and translation were expressed at high levels at 4 and 5 wpi, whereas, at 9 wpi, the Flp pilus genes were highly expressed, suggesting adherence and reduced mobility of the bacteria. The 'Ca. L. solanacearum' genes encoding ATP and C4-dicarboxylate uptake were differentially expressed between the early and late infection stages, suggesting a change in the dependence on different host-derived energy sources. HPE1 effector and salicylate hydroxylase were expressed, presumably to suppress host cell death and salicylic acid-dependent defenses during the infection.[Formula: see text] Copyright © 2021 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Genomic sequence of 'Candidatus Liberibacter solanacearum' haplotype C and its comparison with haplotype A and B genomes.

Haplotypes A and B of 'Candidatus Liberibacter solanacearum' (CLso) are associated with diseases of solanaceous plants, especially Zebra chip disease of potato, and haplotypes C, D and E are associated with symptoms on apiaceous plants. To date, one complete genome of haplotype B and two high quality draft genomes of haplotype A have been obtained for these unculturable bacteria using metagenomics from the psyllid vector Bactericera cockerelli. Here, we present the first genomic sequences obtained for the carrot-associated CLso. These two genomic sequences of haplotype C, FIN114 (1.24 Mbp) and FIN111 (1.20 Mbp), were obtained from carrot psyllids (Trioza apicalis) harboring CLso. Genomic comparisons between the haplotypes A, B and C revealed that the genome organization differs between these haplotypes, due to large inversions and other recombinations. Comparison of protein-coding genes indicated that the core genome of CLso consists of 885 ortholog groups, with the pan-genome consisting of 1327 ortholog groups. Twenty-seven ortholog groups are unique to CLso haplotype C, whilst 11 ortholog groups shared by the haplotypes A and B, are not found in the haplotype C. Some of these ortholog groups that are not part of the core genome may encode functions related to interactions with the different host plant and psyllid species.

Association of "Candidatus Liberibacter solanacearum" with the psyllid, Trioza apicalis (Hemiptera: Triozidae) in Europe.

The psyllid Trioza apicalis Förster (Hemiptera: Triozidae) is a serious pest of carrots, Daucus carota L., in Europe. Carrots exhibiting symptoms of psyllid damage were observed in commercial fields in southern Finland in 2008. Symptoms in affected plants included leaf curling, yellow and purple discoloration of leaves, stunted growth of shoots and roots, and proliferation of secondary roots. Mechanisms by which T. apicalis induces symptoms in plants are not understood, and no plant pathogens have yet been associated with this insect. Given recent association of liberibacter with several crops affected by psyllids, an investigation on whether this bacterium is associated with T. apicalis was conducted. Polymerase chain reaction (PCR) primer pairs OA2/OI2c and LsoF/OI2c, specific for 16S rRNA gene from "Candidatus Liberibacter solanacearum," generated amplicons of 1,168 bp and 1,173 bp, respectively, from DNA extracted from field-collected psyllids (61 and 36.6%, respectively), laboratory-reared psyllids (70 and 33.3%, respectively), field-collected petioles from symptomatic carrots (80 and 55%, respectively), and laboratory-grown carrots (100% for both primer pairs). In contrast, no PCR products were detected in DNA extracted from insect-free plants. The DNA sequences of amplicons of the genes encoding liberibacter 16S rRNA from psyllids and carrots were identical. DNA of the 16S rRNA gene sequences determined from carrots and psyllids were 99.9% identical to analogous sequences of "Ca. L. solanacearum" amplified from several solanaceous crops and the psyllid Bactericera cockerelli (Sulc), a vector of this bacterium. This is the first report of a plant pathogen associated with T. apicalis and the second known psyllid species associated with "Ca. L. solanacearum".

Elevated atmospheric ozone increases concentration of insecticidal Bacillus thuringiensis (Bt) Cry1Ac protein in Bt Brassica napus and reduces feeding of a Bt target herbivore on the non-transgenic parent.

Sustained cultivation of Bacillus thuringiensis (Bt) transgenic crops requires stable transgene expression under variable abiotic conditions. We studied the interactions of Bt toxin production and chronic ozone exposure in Bt cry1Ac-transgenic oilseed rape and found that the insect resistance trait is robust under ozone elevations. Bt Cry1Ac concentrations were higher in the leaves of Bt oilseed rape grown under elevated ozone compared to control treatment, measured either per leaf fresh weight or per total soluble protein of leaves. The mean relative growth rate of a Bt target herbivore, Plutella xylostella L. larvae was negative on Bt plants in all ozone treatments. On the non-transgenic plants, larval feeding damage was reduced under elevated ozone. Our results indicate the need for monitoring fluctuations in Bt toxin concentrations to reveal the potential of ozone exposure for altering dosing of Bt proteins to target and non-target herbivores in field environments experiencing increasing ozone pollution.

Effects of elevated carbon dioxide and ozone on volatile terpenoid emissions and multitrophic communication of transgenic insecticidal oilseed rape (Brassica napus).

Does transgenically incorporated insect resistance affect constitutive and herbivore-inducible terpenoid emissions and multitrophic communication under elevated atmospheric CO(2) or ozone (O(3))? This study aimed to clarify the possible interactions between allocation to direct defences (Bacillus thuringiensis (Bt) toxin production) and that to endogenous indirect defences under future climatic conditions. Terpenoid emissions were measured from vegetative-stage non-Bt and Bt Brassica napus grown in growth chambers under control or doubled CO(2), and control (filtered air) or 100 ppb O(3). The olfactometric orientation of Cotesia vestalis, an endoparasitoid of the herbivorous diamondback moth (Plutella xylostella), was assessed under the corresponding CO(2) and O(3) concentrations. The response of terpenoid emission to CO(2) or O(3) elevations was equivalent for Bt and non-Bt plants, but lower target herbivory reduced herbivore-inducible emissions from Bt plants. Elevated CO(2) increased emissions of most terpenoids, whereas O(3) reduced total terpenoid emissions. Cotesia vestalis orientated to host-damaged plants independent of plant type or CO(2) concentration. Under elevated O(3), host-damaged non-Bt plants attracted 75% of the parasitoids, but only 36.8% of parasitoids orientated to host-damaged Bt plants. Elevated O(3) has the potential to perturb specialized food-web communication in Bt crops.

Constitutive and herbivore-inducible glucosinolate concentrations in oilseed rape (Brassica napus) leaves are not affected by Bt Cry1Ac insertion but change under elevated atmospheric CO2 and O3.

Glucosinolates are plant secondary compounds involved in direct chemical defence by cruciferous plants against herbivores. The glucosinolate profile can be affected by abiotic and biotic environmental stimuli. We studied changes in glucosinolate patterns in leaves of non-transgenic oilseed rape (Brassica napus ssp. oleifera) under elevated atmospheric CO2 or ozone (O3) concentrations and compared them with those from transgenic for herbivore-resistance (Bacillus thuringiensis Cry1Ac endotoxin), to assess herbivory dynamics. Both elevated CO2 and O3 levels decreased indolic glucosinolate concentrations in transgenic and non-transgenic lines, whereas O3 specifically increased the concentration of an aromatic glucosinolate, 2-phenylethylglucosinolate. The herbivore-inducible indolic glucosinolate response was reduced in elevated O3 whereas elevated CO2 altered the induction dynamics of indolic and aliphatic glucosinolates. Herbivore-resistant Bt plants experienced minimal leaf damage after target herbivore Plutella xylostella feeding, but exhibited comparatively similar increase in glucosinolate concentrations after herbivory as non-transgenic plants, indicating that the endogenous glucosinolate defence was not severely compromised by transgenic modifications. The observed differences in constitutive and inducible glucosinolate concentrations of oilseed rape under elevated atmospheric CO2 and O3 might have implications for plant-herbivore interactions in Brassica crop-ecosystems in future climate scenarios.

Influence of carrot psyllid (Trioza apicalis) feeding or exogenous limonene or methyl jasmonate treatment on composition of carrot (Daucus carota) leaf essential oil and headspace volatiles.

The effect of carrot psyllid (Trioza apicalis Förster) feeding and limonene and methyl jasmonate (MeJA) treatments on the essential oil composition and headspace volatiles of carrot (Daucus carota ssp. sativus), cvs. Parano and Splendid, leaves was studied. Carrot psyllid feeding significantly increased the concentrations of sabinene, beta-pinene, and limonene, whereas limonene treatment increased the concentration of (Z)-beta-ocimene in the leaves of both cultivars. The limonene treatment significantly increased the concentration of total phenolics in the leaves of both cultivars, and MeJA treatment increased phenolic concentration in the leaves of Parano. Exogenous limonene spray did not decrease the number of carrot psyllid eggs laid either 2 or 24 h after treatment. The results suggest that carrot psyllid feeding induces changes in the endogenous monoterpene pool in the carrot leaves. Limonene and MeJA treatments affect some induced defenses of the carrot, but the exogenous limonene spray is not an effective oviposition deterrent against carrot psyllid.

Response of Plutella xylostella and its parasitoid Cotesia plutellae to volatile compounds.

The effects of limonene, a mixture of limonene + carvone (1:1, v/v), and methyl jasmonate (MeJA) on diamondback moth (DBM) (Plutella xylostella L.) oviposition, larval feeding, and the behavior of its larval parasitoid Cotesia plutellae (Kurdjumov) with cabbage (Brassica oleracea L. ssp. capitata, cvs. Rinda and Lennox) and broccoli (B. oleracea subsp. Italica cv Lucky) were tested. Limonene showed no deterrent effect on DBM when plants were sprayed with or exposed to limonene, although there was a cultivar difference. A mixture of limonene and carvone released from vermiculite showed a significant repellent effect, reducing the number of eggs laid on the cabbages. MeJA treatment reduced the relative growth rate (RGR) of larvae on cv Lennox leaves. In Y-tube olfactometer tests, C. plutellae preferred the odors of limonene and MeJA to filtered air. In cv Lennox, the parasitoid preferred DBM-damaged plants with limonene to such plants without limonene. C. plutellae females were repelled by the mixture of limonene + carvone. In both cultivars, exogenous MeJA induced the emission of the sesquiterpene (E,E)-alpha-farnesene, the homoterpene (E)-4,8-dimethyl-1,3,7-nonatriene (DMNT), and green leaf volatile (Z)-3-hexenyl acetate + octanal. The attractive effect of limonene and MeJA predicts that these two compounds can be used in sustainable plant protection strategies in organic farming.